RESUMO
KEY MESSAGE: Multiple QTLs control unreduced pollen production in potato. Two major-effect QTLs co-locate with mutant alleles of genes with homology to AtJAS, a known regulator of meiotic spindle orientation. In diploid potato the production of unreduced gametes with a diploid (2n) rather than a haploid (n) number of chromosomes has been widely reported. Besides their evolutionary important role in sexual polyploidisation, unreduced gametes also have a practical value for potato breeding as a bridge between diploid and tetraploid germplasm. Although early articles argued for a monogenic recessive inheritance, the genetic basis of unreduced pollen production in potato has remained elusive. Here, three diploid full-sib populations were genotyped with an amplicon sequencing approach and phenotyped for unreduced pollen production across two growing seasons. We identified two minor-effect and three major-effect QTLs regulating this trait. The two QTLs with the largest effect displayed a recessive inheritance and an additive interaction. Both QTLs co-localised with genes encoding for putative AtJAS homologs, a key regulator of meiosis II spindle orientation in Arabidopsis thaliana. The function of these candidate genes is consistent with the cytological phenotype of mis-oriented metaphase II plates observed in the parental clones. The alleles associated with elevated levels of unreduced pollen showed deleterious mutation events: an exonic transposon insert causing a premature stop, and an amino acid change within a highly conserved domain. Taken together, our findings shed light on the natural variation underlying unreduced pollen production in potato and will facilitate interploidy breeding by enabling marker-assisted selection for this trait.
Assuntos
Arabidopsis , Solanum tuberosum , Melhoramento Vegetal , Pólen/genética , Genótipo , Arabidopsis/genética , MeioseRESUMO
Self-compatible (SC) diploid potatoes allow innovative potato breeding. Therefore, the Sli gene, originally described in S. chacoense, has received much attention. In elite S. tuberosum diploids, spontaneous berry set is occasionally observed. We aimed to map SC from S. tuberosum origin. Two full-sib mapping populations from non-inbred diploids were used. Bulks were composed based on both pollen tube growth and berry set upon selfing. After DNA sequencing of the parents and bulks, we generated k-mer tables. Set algebra and depth filtering were used to identify bulk-specific k-mers. Coupling and repulsion phase k-mers, transmitted from the SC parent, mapped in both populations to the distal end of chromosome 12. Intersection between the k-mers from both populations, in coupling phase with SC, exposed a shared haplotype of approximately 1.5 Mb. Subsequently, we screened read archives of potatoes and wild relatives for k-mers specific to this haplotype. The well-known SC clones US-W4 and RH89-039-16, but surprisingly, also S. chacoense clone M6 were positives. Hence, the S. tuberosum source of SC seems identical to Sli. Furthermore, the candidate region drastically reduced to 333 kb. Haplotype-specific KASP markers were designed and validated on a panel of diploid clones including another renown SC dihaploid G254. Interestingly, k-mers specific to the SC haplotype were common in tetraploid varieties. Pedigree information suggests that the SC haplotype was introduced into tetraploid varieties via the founder "Rough Purple Chili". We show that Sli is surprisingly widespread and indigenous to the cultivated gene pool of potato.
Assuntos
Genética Populacional , Melhoramento Vegetal , Solanum tuberosum/genética , Mapeamento Cromossômico , Diploide , Pool Gênico , Genes de Plantas , Marcadores Genéticos , Genótipo , Haplótipos , Fenótipo , Tubo Polínico/crescimento & desenvolvimento , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Following the molecular characterisation of functional disease resistance genes in recent years, methods to track and verify the integrity of multiple genes in varieties are needed for crop improvement through resistance stacking. Diagnostic resistance gene enrichment sequencing (dRenSeq) enables the high-confidence identification and complete sequence validation of known functional resistance genes in crops. As demonstrated for tetraploid potato varieties, the methodology is more robust and cost-effective in monitoring resistances than whole-genome sequencing and can be used to appraise (trans) gene integrity efficiently. All currently known NB-LRRs effective against viruses, nematodes and the late blight pathogen Phytophthora infestans can be tracked with dRenSeq in potato and hitherto unknown polymorphisms have been identified. The methodology provides a means to improve the speed and efficiency of future disease resistance breeding in crops by directing parental and progeny selection towards effective combinations of resistance genes.
Assuntos
Resistência à Doença/genética , Phytophthora infestans/imunologia , Doenças das Plantas/imunologia , Proteínas de Plantas/genética , Polimorfismo Genético , Solanum tuberosum/genética , Produtos Agrícolas , Melhoramento Vegetal , Doenças das Plantas/parasitologia , Plantas Geneticamente Modificadas , Solanum tuberosum/imunologia , TetraploidiaRESUMO
An important part of the current hazard identification of novel plant varieties is comparative targeted analysis of the novel and reference varieties. Comparative analysis will become much more informative with unbiased analytical approaches, e.g. omics profiling. Data analysis estimating the similarity of new varieties to a reference baseline class of known safe varieties would subsequently greatly facilitate hazard identification. Further biological and eventually toxicological analysis would then only be necessary for varieties that fall outside this reference class. For this purpose, a one-class classifier tool was explored to assess and classify transcriptome profiles of potato (Solanum tuberosum) varieties in a model study. Profiles of six different varieties, two locations of growth, two year of harvest and including biological and technical replication were used to build the model. Two scenarios were applied representing evaluation of a 'different' variety and a 'similar' variety. Within the model higher class distances resulted for the 'different' test set compared with the 'similar' test set. The present study may contribute to a more global hazard identification of novel plant varieties.
Assuntos
Perfilação da Expressão Gênica , Modelos Teóricos , Plantas Geneticamente Modificadas/toxicidade , Solanum tuberosum/genética , TranscriptomaRESUMO
The balanced segregation of homologous chromosomes during meiosis is essential for fertility and is mediated by crossovers (COs). A strong reduction of CO number leads to the unpairing of homologous chromosomes after the withdrawal of the synaptonemal complex. This results in the random segregation of univalents during meiosis I and ultimately to the production of unbalanced and sterile gametes. However, if CO shortage is combined with another meiotic alteration that restitutes the first meiotic division, then uniform and balanced unreduced male gametes, essentially composed of nonrecombinant homologs, are produced. This mitosis-like division is of interest to breeders because it transmits most of the parental heterozygosity to the gametes. In potato, CO shortage, a recessive trait previously referred to as desynapsis, was tentatively mapped to chromosome 8. In this article, we have fine-mapped the position of the CO shortage locus and identified StMSH4, an essential component of the class I CO pathway, as the most likely candidate gene. A 7 base-pair insertion in the second exon of StMSH4 was found to be associated with CO shortage in our mapping population. We also identified a second allele with a 3,820 base-pair insertion and confirmed that both alleles cannot complement each other. Such nonfunctional alleles appear to be common in potato cultivars. More than half of the varieties we tested are carriers of mutational load at the StMSH4 locus. With this new information, breeders can choose to remove alleles associated with CO shortage from their germplasm to improve fertility or to use them to produce highly uniform unreduced male gametes in alternative breeding schemes.
Assuntos
Infertilidade , Solanum tuberosum , Alelos , Solanum tuberosum/genética , Melhoramento Vegetal , Meiose/genética , Pólen/genética , Infertilidade/genéticaRESUMO
We genotyped a population of 618 diploid potato clones derived from six independent potato-breeding programmes from NW-Europe. The diploids were phenotyped for 23 traits, using standardised protocols and common check varieties, enabling us to derive whole population estimators for most traits. We subsequently performed a Genome-Wide Association Study (GWAS) to identify quantitative trait loci (QTL) for all traits with SNPs and short-read haplotypes derived from read-backed phasing. In this study, we used a marker platform called PotatoMASH (Potato Multi-Allele Scanning Haplotags); a pooled multiplex amplicon sequencing based approach. Through this method, neighbouring SNPs within an amplicon can be combined to generate multi-allelic short-read haplotypes (haplotags) that capture recombination history between the constituent SNPs, and reflect the allelic diversity of a given locus in a different way than single bi-allelic SNPs. We found a total of 37 unique QTL across both marker types. A core of 10 QTL were detected with SNPs as well as with haplotags. Haplotags allowed to detect an additional 14 QTL not found based on the SNP set. Conversely, the bi-allelic SNP set also found 13 QTL not detectable using the haplotag set. We conclude that both marker types should routinely be used in parallel to maximize the QTL detection power. We report 19 novel QTL for nine traits: Skin Smoothness, Sprout Dormancy, Total Tuber Number, Tuber Length, Yield, Chipping Colour, After-cooking Blackening, Cooking Type and Eye depth.
RESUMO
We have investigated the genetics and molecular biology of orange flesh colour in potato (Solanum tuberosum L.). To this end the natural diversity in three genes of the carotenoid pathway was assessed by SNP analyses. Association analysis was performed between SNP haplotypes and flesh colour phenotypes in diploid and tetraploid potato genotypes. We observed that among eleven beta-carotene hydroxylase 2 (Chy2) alleles only one dominant allele has a major effect, changing white into yellow flesh colour. In contrast, none of the lycopene epsilon cyclase (Lcye) alleles seemed to have a large effect on flesh colour. Analysis of zeaxanthin epoxidase (Zep) alleles showed that all (diploid) genotypes with orange tuber flesh were homozygous for one specific Zep allele. This Zep allele showed a reduced level of expression. The complete genomic sequence of the recessive Zep allele, including the promoter, was determined, and compared with the sequence of other Zep alleles. The most striking difference was the presence of a non-LTR retrotransposon sequence in intron 1 of the recessive Zep allele, which was absent in all other Zep alleles investigated. We hypothesise that the presence of this large sequence in intron 1 caused the lower expression level, resulting in reduced Zep activity and accumulation of zeaxanthin. Only genotypes combining presence of the dominant Chy2 allele with homozygosity for the recessive Zep allele produced orange-fleshed tubers that accumulated large amounts of zeaxanthin.
Assuntos
Carotenoides/biossíntese , Tubérculos/metabolismo , Solanum tuberosum/metabolismo , Xantofilas/metabolismo , Alelos , Vias Biossintéticas , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Liases Intramoleculares/genética , Liases Intramoleculares/metabolismo , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Oxirredutases/genética , Oxirredutases/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Tubérculos/enzimologia , Tubérculos/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Solanum tuberosum/enzimologia , Solanum tuberosum/genética , ZeaxantinasRESUMO
Despite efforts to control late blight in potatoes by introducing R(pi)-genes from wild species into cultivated potato, there are still concerns regarding the durability and level of resistance. Pyramiding R(pi)-genes can be a solution to increase both durability and level of resistance. In this study, two resistance genes, R(Pi-mcd1) and R(Pi-ber), introgressed from the wild tuber-bearing potato species Solanum microdontum and S. berthaultii were combined in a diploid S. tuberosum population. Individual genotypes from this population were classified after four groups, carrying no R(pi)-gene, with only R (Pi-mcd1), with only R(Pi-ber), and a group with the pyramided R(Pi-mcd1) and R (Pi-ber) by means of tightly linked molecular markers. The levels of resistance between the groups were compared in a field experiment in 2007. The group with R(Pi-mcd1) showed a significant delay to reach 50% infection of the leaf area of 3 days. The group with R ( Pi-ber ) showed a delay of 3 weeks. The resistance level in the pyramid group suggested an additive effect of R (Pi-mcd1) with R(Pi-ber). This suggests that potato breeding can benefit from combining individual R(pi)-genes, irrespective of the weak effect of R(Pi-mcd1) or the strong effect of R(Pi-ber).
Assuntos
Genes de Plantas , Imunidade Inata/genética , Phytophthora infestans/imunologia , Doenças das Plantas , Solanum tuberosum , Cruzamento , Produtos Agrícolas/genética , Produtos Agrícolas/microbiologia , DNA de Plantas/genética , Ligação Genética , Marcadores Genéticos , Dados de Sequência Molecular , Phytophthora infestans/patogenicidade , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Doenças das Plantas/microbiologia , Solanum tuberosum/genética , Solanum tuberosum/microbiologiaRESUMO
Resistance to Globodera pallida Rookmaker (Pa3), originating from wild species Solanum tarijense was identified by QTL analysis and can be largely ascribed to one major QTL. GpaXI ( tar ) ( l ) explained 81.3% of the phenotypic variance in the disease test. GpaXI ( tar ) ( l ) is mapped to the long arm of chromosome 11. Another minor QTL explained 5.3% of the phenotypic variance and mapped to the long arm of chromosome 9. Clones containing both QTL showed no lower cyst counts than clones with only GpaXI ( tar ) ( l ) . After Mendelising the phenotypic data, GpaXI ( tar ) ( l ) could be more precisely mapped near markers GP163 and FEN427, thus anchoring GpaXI ( tar ) ( l ) to a region with a known R-gene cluster containing virus and nematode resistance genes.
Assuntos
Cromossomos de Plantas , Nematoides/fisiologia , Solanum/genética , Animais , Mapeamento Cromossômico , Família Multigênica , Locos de Características QuantitativasRESUMO
The distinction between field resistance and resistance based on resistance (R) genes has been proven valid for many plant-pathogen interactions. This distinction does not seem to be valid for the interaction between potato and late blight. In this study, a locus involved in late blight resistance, derived from Solanum microdontum, provides additional evidence for this lack of distinction. The resistance is associated with a hypersensitive response and results in a delay of infection of approximately 1 to 2 weeks. Both a quantitative as well as a qualitative genetic approach were used, based on data from a field assay. Quantitative trait locus (QTL) analysis identified a QTL on chromosome 4 after correction of the resistance data for plant maturity. A qualitative genetic analysis resulted in the positioning of this locus on the short arm of chromosome 4 in between amplified fragment length polymorphism marker pCTmACG_310 and cleaved amplified polymorphic sequence markers TG339 and T0703. This position coincides with a conserved Phytophthora R gene cluster which includes R2, R(2-like), R(Pi-blb3), and R(Pi-abpt). This implies that R(Pi-mcd1) is the fifth R gene of this nucleotide-binding site leucine-rich repeat cluster. The implications of our results on R-gene-based and field resistance are discussed.
Assuntos
Genes de Plantas , Phytophthora/patogenicidade , Solanum tuberosum/genética , Solanum/genética , Solanum/microbiologia , Sequência de Bases , Mapeamento Cromossômico , Cromossomos de Plantas/genética , DNA de Plantas/genética , Marcadores Genéticos , Interações Hospedeiro-Patógeno/genética , Modelos Genéticos , Família Multigênica , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Locos de Características QuantitativasRESUMO
From biological and genetic standpoints, centromeres play an important role in the delivery of the chromosome complement to the daughter cells at cell division. The positions of the centromeres of potato were determined by half-tetrad analysis in a 4x-2x population where the male parent produced 2n pollen by first-division restitution (FDR). The genetic linkage groups and locations of 95 male parent-derived amplified fragment length polymorphism markers could be determined by comparing their position on a 2x-2x highly saturated linkage map of potato. Ten centromere positions were identified by 100% heterozygosity transmitted from the 2n heterozygous gametes of the paternal parent into the tetraploid offspring. The position of these centromeric marker loci was in accordance with those predicted by the saturated 2x-2x map using the level of marker clustering as a criterion. Two remaining centromere positions could be determined by extrapolation. The frequent observation of transmission of 100% heterozygosity proves that the meiotic restitution mechanism is exclusively based on FDR. Additional investigations on the position of recombination events of three chromosomes with sufficient numbers of markers showed that only one crossover occurred per chromosome arm, proving strong interference of recombination between centromere and telomere.
Assuntos
Centrômero/genética , Posicionamento Cromossômico/genética , Cruzamentos Genéticos , Técnicas Genéticas , Solanum tuberosum/genética , Mapeamento Cromossômico , Segregação de Cromossomos/genética , Cromossomos de Plantas/genética , Marcadores Genéticos , Heterozigoto , Polimorfismo GenéticoRESUMO
Before commercial release, new potato (Solanum tuberosum) varieties must be evaluated for content of toxic compounds such as glycoalkaloids (GAs), which are potent poisons. GA biosynthesis proceeds via the cholesterol pathway to α-chaconine and α-solanine. The goal of this study was to evaluate the relationship between total glycoalkaloid (TGA) content and the expression of GAME, SGT1, and SGT3 genes in potato tubers. TGA content was measured by HPLC-MS, and reverse transcription quantitative polymerase chain reactions were performed to determine the relative expression of GAME, SGT1, and SGT3 genes. We searched for cis-elements of the transcription start site using the PlantPAN database. There was a relationship between TGA content and the relative expression of GAME, SGT1, and SGT3 genes in potato tubers. Putative promoter regions showed the presence of several cis-elements related to biotic and abiotic stresses and light. These findings provide an important step toward understanding TGA regulation and variation in potato tubers.
Assuntos
Alcaloides/biossíntese , Proteínas de Plantas/genética , Solanum tuberosum/genética , Solanum tuberosum/metabolismo , Transcrição Gênica , Alcaloides/toxicidade , Vias Biossintéticas , Proteínas de Plantas/metabolismo , Tubérculos/química , Tubérculos/genética , Tubérculos/metabolismo , Regiões Promotoras Genéticas , Solanina/análogos & derivados , Solanina/metabolismo , Solanina/toxicidadeRESUMO
Potato (Solanum tuberosum) yield has increased dramatically over the last 50 years and this has been achieved by a combination of improved agronomy and biotechnology efforts. Gene studies are taking place to improve new qualities and develop new cultivars. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) is a bench-marking analytical tool for gene expression analysis, but its accuracy is highly dependent on a reliable normalization strategy of an invariant reference genes. For this reason, the goal of this work was to select and validate reference genes for transcriptional analysis of edible tubers of potato. To do so, RT-qPCR primers were designed for ten genes with relatively stable expression in potato tubers as observed in RNA-Seq experiments. Primers were designed across exon boundaries to avoid genomic DNA contamination. Differences were observed in the ranking of candidate genes identified by geNorm, NormFinder and BestKeeper algorithms. The ranks determined by geNorm and NormFinder were very similar and for all samples the most stable candidates were C2, exocyst complex component sec3 (SEC3) and ATCUL3/ATCUL3A/CUL3/CUL3A (CUL3A). According to BestKeeper, the importin alpha and ubiquitin-associated/ts-n genes were the most stable. Three genes were selected as reference genes for potato edible tubers in RT-qPCR studies. The first one, called C2, was selected in common by NormFinder and geNorm, the second one is SEC3, selected by NormFinder, and the third one is CUL3A, selected by geNorm. Appropriate reference genes identified in this work will help to improve the accuracy of gene expression quantification analyses by taking into account differences that may be observed in RNA quality or reverse transcription efficiency across the samples.
Assuntos
Biologia Computacional/métodos , Perfilação da Expressão Gênica/normas , Genes de Plantas/genética , Tubérculos/genética , Solanum tuberosum/genética , Transcrição Gênica , Algoritmos , Variação Genética , Padrões de Referência , Análise de Sequência de RNARESUMO
The R3 locus of potato (Solanum tuberosum L.) confers full resistance to avirulent isolates of Phytophthora infestans, the causal agent of late blight. R3 resides in the distal part of chromosome 11 and segregates in a potato mapping population, from which a well-saturated amplified fragment length polymorphism map is available. Using a population of 1,748 plants, we constructed a high-resolution genetic map at the R3 locus. Using the combination of fine mapping and accurate disease testing with specific P. infestans isolates, we detected that the R3 locus is composed of two genes with distinct specificities. The two genes R3a and R3b are 0.4 cM apart and have both been introgressed from S. demissum, the 'donor' species of most characterized race-specific R genes to P. infestans. A natural recombinant between R3a and R3b was discovered in one accession of S. demissum. The synteny between the R3 locus and the tomato I2 locus is discussed.